Thermoreversibly Assembled Polymersomes for Highly Efficient Loading, Processing, and Delivery of Protein and siRNA Biologics

Abstract

Although great advances in siRNA and mRNA delivery have been made (e.g. with lipid nanoparticles), there remains a pressing need for a versatile platform that can deliver both RNA and protein payloads with convenient processing and storage. Here, we demonstrate a highly efficient approach to forming ca. 100 nm vesicular polymer nanoparticles (polymersomes) that requires no organic solvents by utilizing thermoresponsive block copolymers capable of self-assembling in aqueous medium as the solution is raised to room temperature. This is achieved using block copolymers with a domain that has a lower critical solution temperature (LCST) such that they are soluble in aqueous medium under standard refrigeration (4-7°C) temperature but assemble upon warming to room temperature, resulting in large batches of nanoparticles with predictable size and morphology as dictated by polymer structure. The nanomaterials are designed with charged and biofunctional moieties to drive payload affinity as well as in-vivo targeting, respectively. Both siRNA and protein payloads can be incorporated during warming at higher than 75% loading efficiencies. Payload-polymer suspensions can also be lyophilized into a dry state, allowing for greater hydrolytic stability under mild (4-7°C) refrigeration conditions. This greatly reduces processing and storage requirements, as the powder can simply be reconstituted in cold aqueous medium, then used as described. Here, we selected biomedical applications that did not require removal of unencapsulated payload, bypassing further purification processes and demonstrating a highly scalable approach for formulating nanoparticle-based treatments. Finally, we demonstrate that our system is capable of in-vivo delivery in the contexts of protein subunit vaccination, prophylactic immune tolerance induction, and siRNA interference therapy in cancer. We believe this polymersome platform addresses several challenges to broad translation of nanoparticle delivery of biologics to the clinic, such as scalability, loading efficiency, quality control, and dry storage.